Colorectal cancer is a leading cause of death in the United States. There is very strong evidence that screening for this disease can reduce colorectal cancer mortality. Present screening methods include fecal occult blood testing (FOBT) and endoscopy, which can detect some early colon cancers and large polyps. However, FOBT screening suffers from large numbers of false positive and negative results. Whereas periodic sigmoidoscopy can significantly reduce mortality from colon cancer, currently only about 7 percent of eligible persons receive sigmoidoscopic screening. The goal of this project is to develop novel non-invasive molecular biomarkers for the diagnosis of human colon cancer based on multiplex PCR amplification of mRNA coding for eicosanoid-metabolizing enzymes in fecal matter, using quantitative PCR methodology. The expression of the target genes will be normalized relative to an epithelial and a leukocyte cell marker, cytokeratine-19 and leukocyte , respectively. In preliminary studies, significant levels of Cox-2 mRNA were found in fecal matter of older Min mice, with little or no Cox-2 mRNA in the fecal matter of young Min mice or control wild type mice. Specific aims for Phase I research include: development and optimization of real-time multiplex RT-PCR conditions suitable for isolation and quantification of selected mRNA species in human fecal matter, and preliminary studies on the utility of quantitative multiplex RT-PCR for the diagnosis of human colon cancer using coded fecal samples obtained from normal individuals and from colon cancer patients prior to surgical removal of the colon. PROPOSED COMMERCIAL APPLICATIONS: There is a great need, and a very larger market for a reliable non-invasive diagnostic screening test for human colon cancer. The multiplex RT-PCR quantification of fecal mRNAs which encode selected proteins in eicosanoid metabolism and inflammatory pathways is expected to provide a sensitive and reliable method for the differential diagnosis of human colon cancer.